This proposed research represents an attempt to elucidate the significance of the erythrocyte membrane autophosphorylation system. The human erythrocyte membranes contain both cyclic AMP-dependent and -independent protein kinase activities. Between these two kinases, they catalyze the phosphorylation of several erythrocyte membrane proteins. However, the major membrane polypeptides, spectrin and band 3 (transmembrane protein), are phosphorylated by the cyclic AMP-independent protein kinase. We will focus our attention on the study of this kinase and its action towards spectrin. Attempt will be made to purify the cyclic AMP-independent protein kinase to homogeneity. The kinetic and molecular properties of the kinase will be determined. Antibody against the kinase will be prepared and its interaction with the erythrocyte ghost will be investigated. From the studies on the inhibition of the cyclic AMP-independent protein kinase by its antibody in the membrane autophosphorylation reaction, several useful data can be obtained. It will help us to further clarify the substrate specificities of both the cyclic AMP-dependent and -independent protein kinase within the erythrocyte membranes. It will also permit us to examine the role of these kinases in the maintenance of red cell shape. Finally, it should be possible to determine whether these kinases are altered or absent in certain abnormal erythrocyte membranes. In view of the implication that phosphorylation of spectrin might mediate the ATP-induced red cell shape changes, the phosphorylation of spectrin by the cycle AMP-independent protein kinase will be investigated in greater details. The significance of spectrin phosphorylation in red cell shape changes will be studied by reconstituting erythrocyte ghosts containing the phosphospectrin. This will permit us to examine specifically the effect of phosphospectrin on the shape of the reconstituted ghosts. Finally, attempt will also be made to investigate the effect of phosphorylation of spectrin and of other membrane proteins on cross-linking by the Ca-dependent transglutaminase.